Background for therapeutic use of laminin and peptide fragments of laminin in the treatment of Alzheimer's disease and other amyloidoses can be found in U.S. patent application Ser. No. 09/938,275 filed Aug. 22, 2001, the text and drawings of which are hereby incorporated by reference into the present application as if fully set forth herein.
Beta-Amyloid Protein as a Therapeutic Target for Alzheimer's Disease
Alzheimer's disease (AD) is characterized by the deposition and accumulation of a 39–43 amino acid peptide termed the beta-amyloid protein, Aβ or β/A4 (Glenner and Wong, Biochem. Biophys. Res. Comm. 120:885–890. 1984; Masters et al, Proc. Nat. Acad. Sci. U.S.A. 82:4245–4249, 1985; Husby et al, Bull. WHO 71:105–108, 1993). Aβ is derived from larger precursor proteins termed beta amyloid precursor proteins (or APPs) of which there are several alternatively spliced variants. The most abundant forms of the APPs include proteins consisting of 695, 751 and 770 amino acids (Kitaguchi et al, Nature 331:530–532, 1988; Ponte et al, Nature 331:525–527, 1988; Tanzi et al, Nature 331:528–530, 1988). The small Aβ peptide is a major component that makes up the core of amyloid deposits called “plaques” in the brains of patients with AD. In addition, AD is characterized by the presence of numerous neurofibrillary “tangles”, consisting of paired helical filaments which abnormally accumulate in the neuronal cytoplasm (Grundke-Iqbal et al Proc. Natl. Acad. Sci. U.S.A. 83:4913–4917, 1986; Kosik et al, Proc. Natl. Acad. Sci. U.S.A. 83:4044–4048, 1986; Lee et al, Science 251:675–678, 1991). The other major type of lesion found in AD brain is the accumulation of amyloid in the walls of blood vessels, both within the brain parenchyma and meningeal vessels that lie outside the brain. The amyloid deposits localized to the walls of blood vessels are referred to as cerebrovascular amyloid or congophilic angiopathy (Mandybur, J. Neuropath. Exp. Neurol. 45:79–90, 1986; Pardridge et al, J. Neurochem. 49:1394–1401, 1987). The pathological hallmarks of AD therefore are the presence of “plaques”, “tangles”, and cerebrovascular amyloid deposits.
For many years there has been an ongoing scientific debate as to the importance of “amyloid” in AD and whether the “plaques” and “tangles” characteristic of this disease, were a cause or merely the consequences of the disease. Recent studies indicate that amyloid is indeed a causative factor for AD and should not be regarded merely as a consequence. The Alzheimer's Aβ protein in cell culture has been shown to cause degeneration of nerve cells within a short time period (Pike et al, Br. Res. 563:311–314, 1991; J. Neurochem. 64:253–265, 1995). Studies suggest that it is the fibrillar structure, characteristic of all amyloids, that is mainly responsible for the neurologic effects. Aβ has also been found to be neurologic in slice cultures of hippocampus (Hadrian et al, Neurobiol. Aging 16:779–789, 1995) and induces nerve cell death in transgenic mice (Games et al, Nature 373:523–527, 1995; Hsiao et al, Science 274:99–102, 1996). Injection of Aβ into rat brain also causes memory impairment and neuronal dysfunction (Flood et al, Proc. Natl. Acad. Sci. U.S.A. 88:3363–3366, 1991; Br. Res. 663:271–276, 1994). Convincing evidence that Aβ amyloid is directly involved in the pathogenesis of AD comes from genetic studies. It was discovered that the increased production of Aβ could result from mutations in the gene encoding, its precursor, APP (Van Broeckhoven et al, Science 248:1120–1122, 1990; Murrell et al, Science 254:97–99, 1991; Haass et al, Nature Med. 1:1291–1296, 1995). The identification of mutations in the APP gene which causes early onset familial AD is a strong argument that Aβ and amyloid are central to the pathogenetic process underlying this disease. Four reported disease-causing mutations have now been discovered which demonstrate the importance of Aβ in causing familial AD (reviewed in Hardy, Nature Gen. 1:233–234, 1992). Lastly, recent studies suggest that a reduction in amyloid plaque load in APP transgenic mice lead to improvements in behavioral impairment and memory loss (Chen et al, Nature 408:978–982, 2000; Janus et al, Nature 408:979–982, 2000; Morgan et al, Nature 408:982–985, 2000). This is the strongest argument to date that implicates that reduction of Aβ amyloid load in brain should be a central target for the development of new and effective treatments of AD and related disorders.
Alzheimer's Disease and the Aging Population
Alzheimer's disease is a leading cause of dementia in the elderly, affecting 5–10% of the population over the age of 65 years (Jorm, A Guide to Understanding of Alzheimer's Disease and Related Disorders, New York University Press, New York, 1987). In AD, the parts of the brain essential for cognitive processes such as memory, attention, language, and reasoning degenerate. In some inherited forms of AD, onset is in middle age, but more commonly, symptoms appear from the mid-60's onward. AD today affects 4–5 million Americans, with slightly more than half of these people receiving care in many different health care institutions. The prevalence of AD and other dementias doubles every 5 years beyond the age of 65, and recent studies indicate that nearly 50% of all people age 85 and older have symptoms of AD (NIH Progress Report on AD, National Institute on Aging, 2000). Thirty-three million people of the total population of the United States are age 65 and older, and this will climb to 51 million people by the year 2025 (NIH Progress Report on AD, National Institute on Aging, 2000). The annual economic toll of AD in the United States in terms of health care expenses and lost wages of both patients and their caregivers is estimated at $80 to $100 billion (NIH Progress Report on AD, National Institute on Aging, 2000).
Tacrine hydrochloride (“Cognex”), the first FDA approved drug for AD is an acetylcholinesterase inhibitor (Cutler and Sramek, N. Engl. J. Med. 328:808–810, 1993). However, this drug has showed limited success in the cognitive improvement in AD patients and initially had major side effects such as liver toxicity. The second and third FDA approved drugs for AD, are donepezil (“Aricept”)(Barner and Gray, Ann. Pharmacotherapy 32:70–77, 1998; Rogers and Friedhoff, Eur. Neuropsych. 8:67–75, 1998), and rivastigmine tartrate (“E2020” or “Exelon”) (Polinsky, Clin. Ther. 20:634–647, 1998; Ballard and McAllister, Pychopharmacol. 146:10–18, 1999), which are also acetylcholinesterase inhibitors and more effective than Tacrine in demonstrating slight cognitive improvements in AD patients, but are not believed to be a cure. Therefore, it is clear that there is a need for more effective treatments for AD patients. In the present invention, we have identified laminin globular domain-derived peptides that serve as potent inhibitors of Aβ fibril formation and growth, and which cause disruption/disassembly of preformed AD fibrils.
Laminin and its Presence in Alzheimer's Disease
Laminin is a large glycoprotein complex of 850 kDa which normally resides on the basement membrane and is produced by a variety of cells including embryonic, epithelial and tumor cells (Foidart et al Lab. Invest. 42:336–342, 1980; Timpl, Eur. J. Biochem. 180:487–502, 1989). Laminin interacts with various extracellular matrix components including heparan sulfate proteoglycans (Riopelle and Dow, Br. Res. 525:92–100, 1990; Battaglia et al, Eur. J. Biochem. 208:359–366, 1992), heparin (Sakashita et al, FEBS Letts. 116:243–246, 1980; Del-Rosso et al, Biochem. J. 199:699–704, 1981; Skubitz et al, J. Biol. Chem. 263:4861–4868, 1988) and type IV collagen (Terranova et al, Cell 22:719–726, 1980; Rao et al, Biochem. Biophys. Res. Comm. 128:45–52, 1985; Charonis et al, J. Cell Biol. 100:1848–1853, 1985; Laurie et al, J. Mol. Biol. 189:205–216, 1986). Laminin is composed of three distinct polypeptide chains, A1, B1 and B2 (also referred to as alpha-1, β1 and gamma-1, respectively), joined in a multidomain cruciform structure possessing three short arms and one long arm (Burgeson et al, Matrix Biol. 14:209–211, 1994). Studies involving in vitro self-assembly and the analysis of cell-formed basement membranes have shown that the three short arms interact to form a polymer which is a part of a basement membrane network (Yurchenco et al, J. Biol. Chem. 260:7636–7644, 1985; J. Cell Biol. 117:1119–1133, 1992; Yurchenco and Cheng, J. Biol. Chem. 268:17286–17299, 1993). In addition to its role in basement membrane formation (Kleinman et al, Biochem. 22:4969–4974, 1983), laminin also plays important roles in a number of fundamental biological processes including promotion of neurite outgrowth (Lander et al, Proc. Natl. Acad. Sci. U.S.A. 82:2183–2187, 1985; Bronner-Fraser and Lallier, J. Cell Biol. 106:1321–1329, 1988) and cell adhesion (Engvall et al, J. Cell Biol. 103:2457–2465, 1986). Injury to adult brain also induces laminin production by astrocytes (Liesi et al, EMBO J. 3:683–686, 1984) indicating its role in repair processes. In AD and Down's syndrome, laminin is believed to be present in the vicinity of Aβ amyloid plaques (Perlmutter and Chui, Br. Res. Bull. 24:677–686, 1990; Murtomaki et al, J. Neurosc. Res. 32:261–273, 1992; Perlmutter et al, Micro. Res. Tech. 28:204–215, 1994). Previous studies have also indicated that the various isoforms of APP of AD bind laminin (Narindrasorasak et al, Lab. Invest. 67:643–652, 1992) and other basement membrane components, including perlecan (Narindrasorasak et al, J. Biol. Chem. 266:12878–12883, 1991), fibronectin and type IV collagen (Narindrasorasak et al, J. Biol. Chem. 270:20583–20590, 1995).